Department of Cardiovascular Medicine, The First Affiliated Hospital of Henan Science & Technology University, No. 24, Jinghua Road, Jianxi District, 471003, Luoyang, Henan Province, China.
Departmeng of Urology Surgery, The First Affiliated Hospital of Henan Science & Technology University, No. 24, Jinghua Road, Jianxi District, 471003, Luoyang, Henan Province, China.
Cell Tissue Res. 2021 Sep;385(3):739-751. doi: 10.1007/s00441-021-03463-2. Epub 2021 May 8.
This study analyzed the roles of puerarin and LncRNA ANRIL in myocardial ischemia-reperfusion injury. Hypoxia/reperfusion (H/R) model was established with H9C2 cells. Effects of puerarin of gradient concentrations on cardiomyocytes at different time points of hypoxia and reoxygenation were detected by quantitative real-time polymerase chain reaction (qRT-PCR), cell counting kit-8 (CCK-8), and microscope observation. Effects of puerarin on cardiomyocyte viability, ANRIL expression, contents of lactate dehydrogenase (LDH) and malondialdehyde (MDA), apoptosis, and expressions of autophagy-related genes after H/R injury were determined by CCK-8, quantitative real-time polymerase chain reaction (qRT-PCR), ELISA, flow cytometry, and Western blot, respectively. After cell transfection, the effects of overexpressed and knockdown of ANRIL on cardiomyocytes and H/R-injured cardiomyocytes were examined by rescue experiments. The ischemia-reperfusion (I/R)-injured rat model was established to examine the protective effect of puerarin in vivo. Prolonged hypoxia downregulated ANRIL expression in cardiomyocytes and reduced cardiomyocyte viability. Prolonged reoxygenation increased apoptosis. Both cardiomyocyte viability and ANRIL expression showed a dose-dependent relationship with puerarin. Puerarin reversed the effects of H/R injury on cardiomyocyte viability, ANRIL expression, contents of LDH and MDA, apoptosis, and expressions of autophagy-related genes. Overexpression and knockdown of ANRIL regulated the functions of cardiomyocytes and the expressions of autophagy-related genes. Puerarin reversed the effects of knockdown of ANRIL on H/R-injured cells. The results of In vivo experiments confirmed that puerarin protected myocardial tissues by up-regulating ANRIL and inhibiting autophagy.
本研究分析了葛根素和长链非编码 RNA ANRIL 在心肌缺血再灌注损伤中的作用。用 H9C2 细胞建立缺氧/复氧（H/R）模型。通过实时荧光定量聚合酶链反应（qRT-PCR）、细胞计数试剂盒-8（CCK-8）和显微镜观察检测不同缺氧和复氧时间点不同浓度葛根素对心肌细胞的作用。通过 CCK-8、实时荧光定量聚合酶链反应（qRT-PCR）、ELISA、流式细胞术和 Western blot 分别检测葛根素对 H/R 损伤后心肌细胞活力、ANRIL 表达、乳酸脱氢酶（LDH）和丙二醛（MDA）含量、细胞凋亡以及自噬相关基因表达的影响。转染细胞后，通过挽救实验检测过表达和敲低 ANRIL 对心肌细胞和 H/R 损伤心肌细胞的影响。建立缺血再灌注（I/R）损伤大鼠模型，体内观察葛根素的保护作用。长时间缺氧下调心肌细胞中 ANRIL 的表达，降低心肌细胞活力。长时间复氧增加细胞凋亡。心肌细胞活力和 ANRIL 表达与葛根素呈剂量依赖性。葛根素逆转了 H/R 损伤对心肌细胞活力、ANRIL 表达、LDH 和 MDA 含量、细胞凋亡以及自噬相关基因表达的影响。过表达和敲低 ANRIL 调节心肌细胞的功能和自噬相关基因的表达。葛根素逆转了敲低 ANRIL 对 H/R 损伤细胞的作用。体内实验结果证实，葛根素通过上调 ANRIL 和抑制自噬来保护心肌组织。
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